Effect of prostate-specific membrane antigen on phospho-ERK,growth and migration of prostate cancer LNCaP cells

AIM: To study the expression of prostate-specific membrane antigen (PSMA) on the level of phospho-ERK, cell growth and migration of prostate cancer LNCaP cells.METHODS: The method of silencing PSMA was established by lentivirus-mediated RNAi in our early experiment. The cells were divided into 3 groups.In experimental group, the expression of PSMA in LNCaP cells was stably blocked by lentivirus-mediated RNAi. In negative control group, the cells were transfected with lentivirus-mediated control RNAi (without any interference to PSMA).The normal LNCaP cells served as blank control. The cells in these 3 groups were cultured in both 2 environments: normal medium and medium with PD98059 (an inhibitor of ERK phosphorylation). The phospho-ERK was detected by Western blotting and immunocytochemistry. Furthermore, the growth and migration of the cells were evaluated by MTT and transwell assays,respectively.RESULTS: In normal medium, the expression of phospho-ERK was attenuated in experimental group (P<0.05) and the quantity of "positive" cells was less than those in other 2 groups (P<0.05). Furthermore, the growth curves of the cells showed that the growth ability in experimental group was significantly decreased (P<0.05, after 48 h) and the migration ability in experimental group was reduced (P<0.05). In the inhibitory medium, the cells in all 3 groups expressed phospho-ERK at a lower level. Moreover, the abilities of growth and migration in these 3 groups were poorly displayed. These inhibitory effects on phosphorylation of ERK were similar to the cells in experimental group cultured in normal medium.CONCLUSION: PSMA may play a role in up-regulation of phospho-ERK and it may take an advantage in growth and migration of prostate cancer LNCaP cells.     

石房蛤毒素单克隆抗体的制备及其特性研究

将石房蛤毒素（Saxitoxin,STX）与牛血清白蛋白(BSA)偶联构建完全抗原STX-BSA,免疫BALB/c小鼠,通过杂交瘤技术获得1株稳定分泌抗STX单克隆抗体的杂交瘤细胞株,单抗亚类为IgM。体内诱生法收集腹水单抗,间接ELISA方法测定抗体效价为1∶102400,抗体亲和常数为3.71×106 L/mol。     

结核分枝杆菌CFP10-ESAT6融合蛋白的真核表达及其牛结核病诊断价值评价

本研究旨在利用毕赤酵母(Pichia pastoris)系统融合表达结核分枝杆菌(Mycobacterium tuberculosis,MTB)的培养滤液蛋白10(culture filter protein 10,CFP10)和早期分泌抗原靶6蛋白(earlier secreted antigen target 6,ESAT6),并评价其作为牛结核病外周血γ-干扰素(interferon gamma,IFN-γ)体外释放实验特异性刺激剂来诊断牛结核病的应用潜能.通过PCR扩增cfp1 0-esat6融合基因,构建pPICZα A-(cfp10-esat6)重组质粒,电转化毕赤酵母GSll5,添加甲醇至终浓度1.0％,诱导3d,取培养上清进行SDS-PAGE分析,镍离子金属螯合亲和层析柱纯化目的蛋白,Western blot分析重组蛋白的免疫反应性;取纯化的融合蛋白CFP10-ESAT6作为刺激剂用于牛结核病外周血IFN-γ体外释放实验,评价其牛结核病诊断价值.结果表明,目的蛋白(33kD)被成功分泌表达,该融合蛋白与抗CFP10、ESAT6、组氨酸标签和c-Myc4种单抗均发生特异性反应,具有较好的免疫反应性.165份奶牛外周血样品的IFN-γ检测结果表明,CFP 10-ESAT6融合蛋白与结核菌素纯蛋白衍生物(purified protein derivative,PPD)作为刺激剂,二者阳性符合率为82.3％,阴性符合率为78.8％.本研究利用酵母表达系统成功表达CFP10-ESAT6融合蛋白,并表现出更高的生物学活性,在牛结核病外周血IFN-γ体外释放实验中可作为候选刺激剂,并能克服混合感染导致的PPD漏检问题,从而进一步提高检测的敏感性和特异性.     

马立克氏病病毒(MDV)糖蛋白gB在重组鸡痘病毒中的表达

将马立克氏病病毒gB基因用EcoR I从质粒pUR-MDVEgB中切下。两端补平后，插入到质粒pEFgpt12s的Pst I酶切位点。构建了含有完整的MDVgB基因的转移载体pEFgpt12s-gB。这个载体的特点是，它含有两个启动子，在强的复合启动子Spromoter的下游是插入的gB基因，另一个反向启动子vvP7.5的下游 是标记基因Ecogpt，它的两端是FPV的非必需区。携带gB基因的质粒pEFgpt12s-gB通过磷酸钙的方法转染用282e4株FPV感染3-4h的鸡胚成纤维细胞。通过药物筛选 ，得到含有gB基因的重组病毒。通过PCR、免疫荧光检测，证实了重组病毒中含有gB基因，并且gB糖蛋白也得到了表达。     

酶标抗原Dot－ELISA检测鸭肝炎病毒抗体

酶标抗原Dot－ELISA检测鸭肝炎病毒抗体@杨奎@陈溥言...     

Salmonella abortus ovis (Etude de 112 souches)

The bacteriological examination of 112 strains of Salmonella abortus ovis shows some particular properties of this serotype, and the existence of two different types according to their geographical source.     

Preparation of antigen for the counterimmunoelectrophoretic test for plasmacytosis in mink.

Counterimmunoelectrophoresis as a test method for making the diagnosis of plasmacytosis in mink demands the specific virus antigen. The method for preparation of the antigen according to Cho & Ingram (1972 a, b) with minor modifications is described in details, and results obtained at 62 antigen preparations are presented. In addition an ultrafiltration method is outlined which may be useful as a replacement for ultracentrifugation procedures used in the technique described by Cho & Ingram (1974).     

大肠杆菌新菌毛抗原的研究Ⅱ．菌毛抗原蛋白质特性测定

用保温法和裂解法可以有效提取Ｆ_(１９８７)新菌毛抗原蛋白质；该菌毛蛋白经区带电泳显示主、次两个组分，在０．０６ｍｏｌ／ＬｐＨ８．６巴比妥缓冲液电泳条件下，均向阳极泳动；免疫电泳显示两条沉淀线；其分子量为１９１００Ｄａ；等电点为３．０；由天门冬氨酸、苏氨酸、丝氨酸、谷氨酸、甘氨酸、丙氨酸、缬氨酸、异亮氨酸、亮氨酸、酪氨酸、苯丙氨酸、赖氨酸、组氨酸、精氨酸、脯氨酸、胱氨酸等１６种氨基酸组成。     

接种泰氏锥虫及其抗原使家兔和小鼠产生抗伊氏锥虫的交叉免疫力

用培养的泰氏锥虫制备的死、活两种虫体抗原,接种于家兔和小鼠,一定时间后用伊氏锥虫强毒株攻击,观察其血清抗体的变化及交叉免疫保护能力。试验表明,泰氏锥虫虫体抗原可刺激机体产生较高的体液抗体;免疫动物有一定的抵御伊氏锥虫攻击的能力,与对照组相比,免疫动物血液中虫体出现的时间较迟,最初的数量少,临床症状轻。死虫抗原组的交叉免疫保护能力较活虫抗原组强。     

片形吸虫分泌排泄抗原和虫体抗原的分析

用牛源肝片吸虫（南京）、牛源大片吸虫（广西）和羊源肝片吸虫（实验感染）制备可溶性虫体抗原（BA）和和分泌排泄抗原（ES），以SDS－PAGE电泳分析比较，结果显示，3株虫体可溶性虫体抗原至少有20条以上的主要蛋白条带，分子量集中于10－90KD之间，它们之间无显著差异，而3株分泌排泄抗原蛋白成分较简单，明显可分的条带不超过5条，分子量集中于10－30KD之间，且均拥有26－28KD的蛋白成分，提示该蛋白应为片形吸虫ES抗原的主要免疫成分。